In recent years, a variety of biosensors have been developed using specific catalytic actions of enzymes, and their applications to the clinical and examining field have been attempted. As inspection items and specimens are now increasing, there is a demand for biosensors which permit rapid and accurate measurements.
With a sensor for glucose, for instance, since diabetics are drastically increasing in number, the measurment and control of blood-sugar levels by hitherto employed procedures in which the blood is centrifugated to obtain plasma and subjected to the measurement, takes a relatively long time. Thus, a sensor of the type which enables one to measure it with the whole blood is now required. In the simplest form, there is known a sensor which includes, similar to a urine test paper, a support and a carrier formed on the support and containing an enzyme reacting only with glucose and a dye which undergoes a change at the time of the enzyme reaction or with a product formed during the enzyme reaction. The blood is added to the carrier, and a change of the dye after a certain period of time is measured visually or optically. However, this system is disadvantageous in that colored matters in the blood greatly interfere with the measurement, resulting in a low accuracy.
In order to overcome the above, there has been proposed a multi-layered carrier for analysis as is particularly shown in FIG. 1 (Japanese Laid-Open Utility Model Application No. 54-178495). This carrier includes a transparent support 1, on which are superposed a reagent layer 2, a developing layer 3, a waterproof layer 4 and a filtration layer 5 in this order. When a blood sample is dropped from the above, solid matters in the blood such as red blood cells, platelets and the like, are initially removed by means of the filtration layer 5, followed by uniform infiltration from a small hole 6 of the waterproof layer 4 toward the developing layer 3. At the reagent layer 2, the reaction is caused to proceed. After completion of the reaction, light is applied from the direction of the arrow through the transparent support 1 to measure a substrate concentration by spectroscopy. As compared with the known simple stick carrier, this carrier has a complicated structure but the removal of blood cells is more improved. However, it takes a long time for the infiltration and the reaction of the blood corpuscles, so that the waterproof layer 4 for preventing the sample from drying is essential, with an attendant problem that the equipment and the carrier become complicated.
On the other hand , a biosensor of the type as shown in FIG. 2 has been proposed (for example, in Japanese Laid-open Patent Application No. 59-166852) as a system in which a specific component in a biological sample is quantitatively determined in high accuracy without resorting to any procedures such as dilution of a liquid sample and agitation. The biosensor includes an insulative substrate 7, a measuring electrode 8 and a counter electrode 9, which have, respectively, leads 10, 11, embedded in the substrate 7, and a perforated body 12 which covers exposed portions of these electrodes and carries an oxydo-reductase and an electron acceptor. When the liquid sample is dropped on the perforated body, whereupon the oxydo-reductase and the electron acceptor in the perforated boy dissolve in the sample liquid, thus permitting the enzyme reaction with a substrate in the liquid sample to proceed. As a result, the electron acceptor is reduced. After completion of the enzyme reaction, the reduced electron acceptor is electrochemically oxidized and a concentration of the substrate in the liquid sample is determined from a current for the oxidation.
In this arrangement, although the measurement can be performed simply by replacing the perforated body every measurement, additional procedures, such as washing, are undesirable for the electrode system. If parts including the electrodes could be disposably replaced whenever the measurement is effected, the measuring product would become very simple but the system would be very expensive in view of the electrode materials such as platinum and the arrangement. Although the platinum electrodes may be formed by a sputtering method or a vacuum evaporation method, they are still expensive in view of the manufacture.
In order to measure a specific component in a biological sample such as blood simply, rapidly and in high accuracy, a desirable type of sensor is one which can yield measurements only by addition of a liquid sample to the sensor without dilution or weighing. In addition, a sensor of the disposable type is also desirable which does not involve any procedures such as washing, sampling and the like.